The broad goal of this project is to contribute to the understanding of the mechanism of blood clot dissolution or fibrinolysis by plasmin. Specifically we propose to determine the covalent structure of the heavy chain of human plasmin. Human plasmin is a high molecular weight protein consisting of two chains linked by a single disulfide bridge. The light chain (MW 25,700) resembles the protease trypsin in its primary structure and function, and contains the catalytic site of plasmin. On the other hand, virtually nothing is known about the structure and function of the heavy chain (MW 48,800), except that it is essential for plasmin activity. Numerous similarities in gross structure and mode of action exist between plasmin and other coagulation proteins, and one can logically ask whether there are similarities in primary structure between plasmin and these or other proteins such as immunoglobulins. Also, does the heavy chain contain the binding sites for its substrate, fibrin, or for the enzymes which activate plasminogen? These questions can be answered in part by determining the amino acid sequence of plasmin and comparing it with other proteins which are currently being sequenced elsewhere. Sequence studies will be carried out using classical methods for protein fragmentation and separation as well as some new inovations, the object being to obtain peptides of a size (15-25 residues) suitable for sequencing by Edman degradation using the automatic solid-phase peptide sequencer, which was developed in our laboratory. This new technique has not yet been widely applied. A second objective of this proposal is to test the solid-phase sequencing method with a large and complex protein, in this case plasmin.